Cyclosporine (CsA) is a potent immunosuppressive agent whose widespread usage is severely restricted by its nephrotoxic side effects. The mechanism by which CsA causes nephrotoxicity is not known, largely because there does not exist a good animal model in which to study the phenomenon. We are hypothesizing that nephrotoxicity is caused by the presence in the blood of "polar" cyclosporine metabolites, rather than the parent compound. We have found that the appearance of these metabolites correlates with human nephrotoxicity, but that rats do not produce these metabolites even when treated with 75 mg/kg/day of CsA. The long-term objectives of this project are to elucidate the role of CsA metabolism in nephrotoxicity and immunosuppression in order to develop a therapeutic regimen that prevents renal allograft rejection without toxicity. The specific aims of their project are: to evaluate the relationship between CsA metabolism and nephrotoxicity in humans and rats; to determine the effect of methylprednisolone, azathioprine, and inhibitors and inducers of the cyt-P450 system on CsA metabolism and nephrotoxicity in the rat; to determine if CsA and the active metabolites synergize with other immunosuppressive agents, including steroids, 6-mercaptopurine, 2-deoxycoformycin, chlorpromazine, colchicine, and cytochalasins A and B; and to determine if the effects of CsA and the active metabolites on Na+/H+ antiport activity and the rise in intracellular free Ca2+ correlate with inhibition of IL-2 production by human T lymphocytes. CsA metabolism will be monitored using high performance liquid chromatography and radio-immunoassay analysis of blood samples. Synergism between CsA and other agents will be determined by measuring their effects on in vitro human T cell activation (increase in pHi and Ca2+i), function (IL-2 production); growth (DNA synthesis and mitosis), and differentiation (generation of cytotoxic T cells) and B cell growth ( DNA synthesis) and differentiation (IgM and IgG synthesis).